Drug Administration Methods
We utilize multiple different methods of drug delivery in our rodent models depending on the research question being asked. These methods range from daily subcutaneous or intraperitoneal injections, surgically implanted osmotic minipumps (pictured) to administer a controlled dose across 28 days, and catheters surgically implanted into the external jugular vein to allow for drug self-administration in the operant chamber.
Operant Chamber
Equipped with a food delivery system and a drug delivery system, all 24 of our operant chambers (MedAssociates) are capable of running food self-administration and drug self-administration testing. For both methods of testing, levers are utilized to give the rat the option of dispensing either a food pellet or drug infusion at their discretion. Data gathered from these chambers help us answer questions about motivation and reward.
Ultrasonic Vocalization (USV) Recording and DeepSqueak Analysis
To measure developmental disruption following maternal opioid self-administration, we assess disruptions in offspring USVs. Recorded in a sound-attenuating chamber (Noldus Ultravox XT), pup calls are analyzed with the MATLAB program DeepSqueak, using neural networks trained to detect USVs, transform them into spectograms, and quantify the number and curvature of the calls.
RT-qPCR
Quantitative reverse transcription PCR is used within our lab to determine how drug exposure changes patterns of gene expression in discrete brain regions, as well as cells grown in culture.
ELISA
The lab performs ELISA assays with blood plasma and cell culture samples to quantify the levels of proteins of interest and how their expression is affected by drug exposure.
Immunofluorescence
The lab performs immunohistochemical staining and fluorescent imaging in cryosectioned brain tissue and cells grown in culture to visualize nuclear, somatic, and transmembrane protein expression in neural and glial populations with regional specificity.
In Situ Hybridization
To acquire spatial resolution of gene expression throughout the brain, we use RNAscope in situ hybridization. With this state-of-the-art multiplex assay, we can visualize up to four target probes within a single sample with exceptional sensitivity.
Cell Culture
We utilize sex-specific primary neuronal cell cultures of the cortex, hippocampus, and striatum to better understand early neurodevelopmental changes within our multi- and transgenerational models. We utilize sholl analysis to determine dendritic complexity with and without continued drug exposure. We can also measure gene and protein expression from these neurons at various time points to understand how prenatal or preconception drug exposure changes neuronal functioning.
Image Analysis
We use a wide variety of tools to quantify protein and gene expression in tissue sections. One such program is Pipsqueak Pro (Rewire Neuro) for AI-assisted cell detection and intensity quantification. Originally developed specifically for analysis of perineuronal nets and the interneurons they enwrap, Pipsqueak Pro has now been trained to detect a wide variety of biomarkers that allows for optimal detection and colocalization capabilities.
Byrnes-Vassoler Lab 